Laminin-521

Laminin-521

Human recombinant laminin-521


Recreating the natural cell niche is key to successful cultivation. The physical, topological, and biochemical expression of the different laminin isoforms in the BM is heterogeneous and tissue specific. 

Laminin-521 (LN-521) is the natural laminin for pluripotent stem cells and therefore reliably facilitates self-renewal of human ES and iPS cells in a chemically defined, feeder-free and xeno-free stem cell culture system. LN-521 enables efficient single-cell passaging of genetically stable and pluripotent stem cells without need of any apoptosis inhibitors for superior quality of your cells and studies. 

After birth, α5-laminins represent the most common laminins in the body and LN-521 supports many diverse tissue cell types, such as cells from pancreas, vascular, nervous and muscular systems.

BUY ONLINE!

Amount
Size
Price
Oty
100µg 1 x 1ml €45
500µg 5 ml €200
1mg 10 x 1ml €420
For larger quantities contact us

Estimated shipping time
1-5 working days

SAVE MONEY

Buy larger quantities and save shipping cost.

SIZE GUIDE

Not sure how much laminin you need? To make it easy, we have created a tool where you can calculate the amount needed for your experiments. Just choose culture well format and fill in the desired coating concentration to see the amount required.

FORMAT
CONC. (ug/cm2).
NO OF WELLS
AMOUNT (uq)

LN-521 STEM CELL MATRIX

Laminin-521 (LN-521) is a key cell adhesion protein of the natural stem cell niche, expressed and secreted by hPSCs in the inner cell mass of the embryo. LN-521 recreates the biologically relevant milieu in vitro and therefore supports robust expansion of human ES and iPS cells. 


The LN-521 stem cell substrate enables high expansion rate, monolayer homogenous growth, and maintained pluripotency. Cells are genetically stable without karyotypic abnormalities, showed with both karyotyping and SNP arrays. 


hESC and iPSC grow faster on LN-521 compared to other feeder-free matrices, facilitating automation. Once adapted to the LN-521 matrix, hPSCs can routinely be cultured as single cells without the addition of ROCKi. LN-521 also supports efficient clonal survival and expansion of hES and iPS cells, greatly facilitating the study of genomic alterations and manipulations in pluripotent cells. Furthermore, renders more efficient reprogramming of specific tissue cells to iPSCs.


The robust support of pluripotent stem cells by laminin-521 enables maintenance of hPSC without weekend feeding.


After birth, LN-521 also represent one of the most common laminins in the body and together with the other laminin isoforms, it supports expansion and differentiation of many diverse tissue cell types. Read more about different laminin cell culture applications.


Download the cell culture applications poster.

KEY BENEFITS OF LN-521 FOR hPSC CULTURE:

  • Defined and xeno-free cell culture matrix - for control and clinical compliance

  • No lot-to-lot variability - more reliable and standardized experiments

  • Homogenous hPSC cultures that are pluripotency and genetically stabile - no need to remove differentiated cell areas

  • High clonal survival - optimal for derivation and gene editing 

  • Triggers authentic cell signalling pathways - provides hPSCs cultures with a more uniform gene expression profile

  • High expansion rate and controlled single-cell passaging - facilitates automation and high-throughput processes

  • Authentic milieu that primes the hPSCs - more efficient differentiation and enhanced cell maturation, polarization and organization

  • Flexible culture system - easy to control and adaptable to any protocol

  • Support weekend feeding - no more weekend work!


Laminin-521 cell culture application note

biorelevant stem cell culture

Human stem cell culture

Easy culturing of human ES and iPS cells in a 
biorelevant and xeno-free environment 

effective clonal culture

Clonal cell culture

Efficient clonal culture and derivation of
human pluripotent stem cells 

Biological relevance

 natural substrate for stem cell culture

Laminin-521 (LN-521) is a key cell adhesion protein of the natural stem cell niche, expressed and secreted by hPSCs in the inner cell mass of the embryo. LN-521 recreates the biologically relevant milieu in vitro and therefore supports robust expansion of human ES and iPS cells. 

  

 


 

Cost-effective stem cell culturing

 Laminin-521 cost effective stem cell culture

The specific LN-521 binding of the α6β1 integrin enables high cell survival and self-renewal and hPSCs grow faster on LN-521 compared to other feeder-free matrices. Due to faster growth rate and higher cell yield, the total cost (medium and matrix) per cell for an average passage is lowest for LN-521. Read more about cost-effective stem cell culture



 

Easy and flexible culture protocols

easy and flexible cell culture protocols

The single-cell split is easy, fast and forgiving, making anyone suitable to culture hPSCs. When using LN-521, your cell culture protocol can easily be made totally defined and xeno-free with your choice of culture medium and dissociation reagent. For reduced labor and cost, the LN-521 matrix supports weekend-free feeding.

  1. Coat plates with LN-521.

  2. Wash the cells with PBS and add dissociation reagent of choice - incubate

  3. Centrifuge the single-cell suspension and resuspend the pellet in fresh medium of choice

  4. Seed as single-cell suspension on fresh LN-521 coated plates


 

Long-term efficient expansion

Rapid and robust stem cell expansion

hESC and iPSC grow faster on LN-521 compared to other feeder-free matrices, facilitating automation. The specific LN-521 binding of the α6β1 integrin enables high cell survival and self-renewal. Cells expand quickly and reach 100% confluency after only 4 days after a 1:10 split. You will thus have 10.000 fold more cells after only 4 passages, making the cell quantities of low-passage hPSCs needed for clinical applications possible.


 

Genetic stability 

genetically stable stem cell culture
 

With LN-521 hPSCs can be seeded as single cells at low densities and cultured long-term without introduction of genetic abnormalities. Cells have been kept on LN-521 for more than 130 passages with stable karyotypes. Importantly, when LN-521 is used for the derivation of new hESC lines, the embryo is not destroyed and the lines originating from a single blastomer are genetically stable.


 

Homogenous hPSC monolayer 

homogenous stem cell culture without spontaneous differentiation

PSCs plated as single-cell suspension on LN-521 grow as a homogenous monolayer without any abnormal genetic aberrations. The specific LN-521 binding of the α6β1 integrin enables high cell survival and self-renewal and once adapted to the LN-521 matrix, hPSCs can routinely be cultured as single cells without the addition of ROCKi. LN-521 also support clonal survival and is a good substrate for derivation and gene editing.  


 

Less labor and complexity

 

 

stem cell culture without spontaneous differentiation 

 

hPSCs can be cultured to near confluence on LN-521 without spontaneous differentiation. The cells remain pluripotent (Oct4+; pink) on LN-521 with no sopntaneous differentiation and there is no need to remove differentiated cell areas (only DAPI; blue) as compared to cells cultured on other substrates.



 

Read the full-length Nature Communications article on biorelevant and effective stem cell culture

COATING PLATES

See instructions for coating protocol, recommended coating volumes and concentrations.


hPSC CULTURE ON LN-521

See instructions for stem cell culture on laminin-521, including:
- transfer
- passaging
- thawing
- cryopreservation of hPSCs on LN-521. 


521-To-Go™ pre-coated plates

When using 521-To-Go™ pre-coated plates, follow the instructions here.


Embryoid body formation

Protocol for embryoid body formation from hPSCs cultured on LN-521

 

 

IMPORTANT NOTES

  • All procedures should be done under sterile conditions using aseptic techniques

  • Avoid long exposure of the protein to ambient temperatures

  • Repeated freeze/thaw should be avoided

  • The laminin stock solution is stable for 3 years when stored at -20°C

  • Thawed, undiluted laminin stock is stable for at least 3 months when stored at +2°C to +8°C under aseptic conditions

  • For your convenience, the coated plates can be kept for up to 4 weeks when stored aseptically at +2°C to +8°C

  • The protocols can easily be made totally defined and xeno-free with your choice of culture medium and dissociation reagent

  • It is important that the cells transferred to the LN-521 matrix are of high quality

  • Some hPSC lines transferred to the LN-521 matrix, might require an adaptation period before they can be cultured according to the single-cell passaging protocol

  • Once adapted to the LN-521 matrix, hPSCs can routinely be cultured as single cells without ROCKi

  • The LN-521 matrix facilitates long-term self-renewal of hPSC without weekend feeding. For reduced labor and cost, follow the reduced feed protocol.

 

Troubleshooting

Laminin plate coating

An uneven cell spread, it’s often a coating issue and could be caused by the following:

  • To low coating concentration used. Increase the coating concentration is high enough to support an even cell growth.
  • DPBS Ca--/Mg-- has been used. We recommend to use DPBS with Ca2+ and Mg2+ since divalent cations are important for the protein structure and function. 
  • NUNC-plates was used. Most plastic work well for laminin coating but we know the laminin coating is not compatible with some NUNC-plates. SARSDET and Corning plate usually work well.
  • Bad coating coverage/the plate has dried out. Ensure that the entire surface is covered by the laminin coating solution when preparing fresh plates. Also, do not let the plate dry out as this will inactivate the laminin coating. Too long time in the incubator or long storage without sealing could cause too much evaporation so that part of the plate dries out (often center).

 
Transfer from other substrates

  • We recommend to transfer the cells as single cells (or as small clumps) and always with the addition of ROCKi for the first few (3-5) passages. Once the cells are adapted to the LN-521 matrix, the cells can be cultured as single cells without ROCKi. This may take up to 5 passages.
  • If the cells are hard to adapt, try increasing the coating concentration to 10 ug/ml and seed at a higher cell density 50,000 – 100,1000 cells/cm2. Once the cells are adapted a lower coating and seeding concentration often can be used.
  • It is important that the cells transferred to the LN-521 matrix are of high quality. LN-521 will generally also support differentiated cells so carefully select only undifferentiated cell areas for transfer. 
  • LN-521 work well in combination with most commercial media brands. However, it is to be expected that cell morphology will look different dependent on the medium used for culture. 


hPSC splitting and seeding

  • We recommend single-cell passage or passage as small clumps. 
  • Stem cells are sensitive and when using enzyme, do not treat the cells too long as that will damage the cells. Cells attach tighter on laminin compared to other matrices and scraping or pipetting without first loose up cells can affect cell integrity and viability which could result in less attachment next day. Less confluent cells need shorter treatment time whereas more confluent cells might need longer treatment time. The cells should detach easily without too much pipetting needed. Do not use too much mechanical force (extensive pipetting or scraping) as that will damage the cells. Increase the dissociation reagent incubation time rather than increasing the force.  If the cells stick very hard to the LN-521 surface, try to lower the coating concentration.
  • After seeding: most cells should have attached after 1 hour and the cells should be evenly distributed over the entire plate. If there is a lot of cell death after seeding, the cells have most likely been treated too harsh during splitting.
  • The day after seeding: the cell has migrated and should have formed small colonies and will continue to expand as a homogenous monolayer. Cells cultured on the LN-521 matrix are ready to be passaged when cell culture is 60-99% confluent. Depending on the cell line, seeding density and on the medium used, cultures are usually passaged 3-6 days after seeding.

Product name

LN521

 

Product use

Cell culture

 

Product application

Human embryonic and induced pluripotent stem cells


 

Documents

MSDS

Storage

-20°C

 

Stability

Three years from date of receipt

 

Stock concentration

0.1mg/ml

 

Use concentration

0.5-2 ug/cm2
Optimal concentration must be determined by the end user

Protein name

Laminin-521 (LN-521)

 

Synonyms

Laminin-11

 

Expressed in vivo

Expressed and secreted by human pluripotent stem cells and can, in addition, also be found in the kidneys, neuromuscular junctions, lungs and placenta.

Clonal culturing of human embryonic stem cells on laminin-521/E-cadherin matrix in defined and xeno-free environment.

Sergey Rodin, Liselotte Antonsson, Colin Niaudet, Oscar E. Simonson, Elina Salmela, Emil M. Hansson, Anna Domogatskaya, Zhijie Xiao, Pauliina Damdimopoulou, Mona Sheikhi, Jose Inzunza, Ann-Sofie Nilsson, Duncan Baker, Raoul Kuiper, Yi Sun, Elisabeth Blennow, Magnus Nordenskjold, Karl-Henrik Grinnemo, Juha Kere, Christer Betsholtz, Outi Hovatta and Karl Tryggvason.

Nature Communications, 2014

This article provides scientific evidence that LN521™ is the optimal matrix for generation and culture of human pluripotent stem cells. It is described in detail how this physiologically relevant laminin establishes genetically stable hESC lines in an efficient, defined, xeno-free and feeder-free procedure, suitable for stem cell banking and regenerative medicine applications.

 

A defined xeno-free and feeder-free culture system for the derivation, expansion and direct differentiation of transgene-free patient-specific induced pluripotent stem cells.

Hong Fang Lu, Chou Chai, Tze Chiun Lim, Meng Fatt Leong, Jia Kai Lim, Shujun Gao, Kah Leong Lim, Andrew C.A. Wan.

Biomaterials, 2014

This article demonstrates LN521 as an optimal defined, xeno- and feeder-free matrix for the reprogramming of human iPS cells. LN521 achieves high-efficiency reprogramming in different media (E8, Nutristem, etc), fast and easy expansion as well as direct differentiation to dopaminergic neurons on LN521. The authors conclude that the efficient transgene-free hiPSC derivation and expansion on LN521 enables clinical applications useful for human patient iPSCs and derivatives for cellular therapy.

 

Optimization of slow cooling cryopreservation for human pluripotent stem cells.

Takamichi Miyazaki, Norio Nakatsuji and Hirofumi Suemori.

Genesis, 2013

This is one of the first customer publications that demonstrates LN521 as an optimal xeno- and feeder-free matrix for pluripotent stem cells. The authors show cells should be cryopreserved as single cells for highest survival which is specifically supported by LN521 that promotes adhesion and self-renewal of fully dissociated single cells in the absence of ROCK inhibitor. They demonstrate 80-90% survival of hPSCs post-thawing and 60% survival following subculture on LN521, allowing for efficient and easy handling of cells and bulk storage of high-quality hPSCs.

Live visualization of chromatin dynamics with fluorescent TALENs.

Yusuke Miyanari, Céline Ziegler-Birling & Maria-Elena Torres-Padilla.

Nature structural & molecular biology, 2013

The authors of this study used LN511 as a coating substrate for mouse embryonic stem cells on glass in order to aquire better images for the study of endogenous repetitive genomic sequences visualized by TALE integration.

 

Efficient and Scalable Expansion of Human Pluripotent Stem Cells Under Clinically Compliant Settings: A View in 2013.

Ying Wang, Linzhao Cheng and Sharon Gerecht.

Annals of Biomedical Engineering, 2013

The authors of this important review highlight the capacity of LN521 and LN511 to successfully support long-term expansion of pluripotent stem cells.

 

Xeno-free culture of human pluripotent stem cells.

Bergström R, Ström S, Holm F, Feki A, Hovatta O.

Methods Mol Biol, 2011

A book chapter describing four different animal-protein free culture systems that are proven efficient in expanding pluripotent human ES cell populations. Two of the systems are with LN511 as substrate but with different medium, and the other two substrates are CELLstart and human foreskin fibroblasts. Control of ground-state pluripotency by allelic regulation of Nanog Miyanari Y, Torres-Padilla ME Nature, 2012 The authors of this study used LN511 as a coating substrate for mouse embryonic stem cells to investigate the role of Nanog for ground-state pluripotency during development. By using LN511 they were able to culture the mouse embryonic stem cells as monolayers and importantly without the use of LIF in a completely defined cell culture environment.